Biological and solid-state nanopores provide a means of sensing biomolecular analytes at the single molecule level. Individual nanopores are typically embedded in thin insulating membranes, providing the only conduit for ionic current to pass between two liquid reservoirs. Utilizing the principles of larger-scale Coulter counters, nanopore experiments relate changes in ionic current to determine the length, size, charge and conformation of charged biomolecules as they are electrophoretically driven through a nanopore in the presence of an external electric field.
While biological nanopores such as α-hemolysin typically offer greater sensitivity and low-noise properties, the supporting lipid bilayer is fragile and of fixed size, limiting their applicability. Solid-state nanopores, on the other hand, are typically fabricated in thin (10-50 nm) insulating membranes, such as silicon nitride or silicon oxide membranes, and can be made of different sizes, be readily integrated with wafer-scale technologies, and are more robust, allowing for a wider range of experimental conditions. Despite these advantages, solid-state nanopore technologies suffer from several practical drawbacks that limit their usefulness for biomolecular studies. While control of nanopore size is possible, it is typically expensive and laborious to achieve, requiring specialized equipment and skilled personnel. For example, nanopores drilled by focused-ion beam have been recently shown to shrink under specific experimental conditions in a scanning electron microscope (SEM). In other approaches, nanopores drilled by transmission electron microscopy (TEM) can expand or shrink depending on the beam conditions and subsequent exposure to aqueous solvents. In these cases, the achievable range of nanopore sizes is limited, difficult to control and even unreliable as the size of the nanopore can change following chemical treatment or when immersed in a particular liquid environment.
The ionic current through solid-state nanopores can also suffer from a high degree of noise, the sources of which are an intensely investigated topic in nanopore literature. While various methods have been proposed to reduce electrical noise, the yield of reliable, stable low-noise nanopores is typically still quite low. Deposition of carbonaceous residues during drilling and imaging can have detrimental effects on the electrical signal quality, often making complete wetting a challenge and causing the formation of nanobubbles that can be difficult to remove. Furthermore, clogging of the nanopore by analyte molecules degrades signal quality rendering pores unusable for further experiment. These effects greatly reduce yield of functional nanopore devices and increase the cost associated with solid-state nanopore research. Thus, reproducible fabrication and tuning of reliable nanopores remains a challenge not only for the academic research environment but for the commercialization of any nanopore-based technology.
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